Apparently normal rat liver epithelial cells (WB-F344) have been widely used in studies pertaining to carcinogenesis. Ionizing radiation, a well known carcinogen, is known to perturb cell-cycle progression in a dose-dependent manner, thereby causing delay in cell proliferation. However, for WB-F344 cells, there is a paucity of such data, which are of substantial importance in understanding their radiation response. Here, the distribution of phases in the cell-cycle and the proliferation ability of WB-F344 cells are characterized at various time points after the cells have been irradiated with different doses of γ-rays.
After WB-F344 cells reached 100% confluence, they were trypsinized and suspended at 3.5 × 105 cells/ml in culture medium. Cells were irradiated in suspension with 137Cs γ-rays at doses from 1–10 Gy. After irradiation, 1 × 105 cells were plated into 60 × 15-mm culture dishes and incubated at 37°C, with 2% CO2 and 98% air. At 12, 24, 36, 48, and 60 h postirradiation, cells were harvested, counted, and subjected to flow cytometric cell-cycle analysis.
Growth curves of WB-F344 cells irradiated with γ-rays started to separate at 36 h postirradiation. By 60 h postirradiation, the growth curves for each of the 10 absorbed doses were distinctly separated. Drastic redistributions of control and irradiated cells within G0/G1-, S-, and G2/M-phases of the cell cycle were observed during the first 36 h of cell growth. At each time point postirradiation, cell-cycle phase profiles of irradiated cells were altered in a dose-dependent manner. In general, there was a strong correlation between the percentage of G2/M-phase cells and absorbed dose, with the exception of 24 h postirradiation. The percentage of G2/M-phase cells increased as a function of time postirradiation, suggestive of delays in the passage of cells through the G2 cell-cycle checkpoint.
This work provides a general description of cell cycle redistribution and repopulation kinetics of WB-F344 cells at various times postirradiation of quiescent cells that were subsequently allowed to proliferate. In general, growth inhibition and delays in progression through G2/M-phase correlated well with radiation dose. These data should be of considerable significance in the design of experiments that examine the radiation response of these cells. © 2004 Wiley-Liss, Inc.